Use of low molecular weight thrombin inhibitors in cholesterol-lowering therapy

ABSTRACT

According to the invention there is provided a cholesterol lowering therapy method and a method for modification of lipid (triglyceride), lipoprotein, and apolipoprotein profiles associated with an increased risk of cardiovascular complications comprising the administration of a low molecular weight thrombin inhibitor, or a pharmaceutically acceptable derivative thereof, to a patient in need thereof.

FIELD OF THE INVENTION

This invention relates to a new use of low molecular weight thrombininhibitors.

BACKGROUND AND PRIOR ART

It is well known that high levels of cholesterol are associated withheart disease. More than half of all US citizens are understood to havecholesterol levels that exceed those recommended, and one in five hascholesterol levels that are considered high.

Cholesterol is involved in the production and maintenance of cellmembranes, as well as the production of sex hormones (includingprogesterone, testosterone, estradiol and cortisol), bile salts andVitamin D. It is formed primarily in the liver, but also in other partsof the body, such as the small intestine.

In healthy individuals, all of the cholesterol that is needed to performthe above-mentioned functions is produced naturally. However, in atypical blood test, of the amount of cholesterol circulating in blood,about 85% is endogenous, the other 15% arising from external sources.Dietary cholesterol usually originates from meat, poultry, fish, seafoodand dairy products. In this respect, high consumption levels of thesefoodstuffs may give rise to increased cholesterol levels in thebloodstream.

Increased cholesterol levels in serum have been associated withatherosclerosis, which is known to increase significantly the risk ofblood vessel blockage (stenosis), and thus the likelihood of anginapectoris, myocardial infarction and other cardiovascular complications,such as stroke.

Cholesterol is insoluble in aqueous environments and thus needs to betransported within the bloodstream by apolipoproteins (Apos). Whenapolipoproteins are associated with cholesterol, complexes known aslipoproteins are formed. The density of these lipoproteins is determinedby the amount of protein in the molecule and, in this respect,low-density lipoproteins (LDLs), which are the major cholesterol carrierin the blood, are known to have more of the negative effects mentionedherein than protective high-density lipoproteins (HDLs). High levels ofLDLs are thus associated with atherosclerosis, whereas greater levels ofHDLs are understood to provide some protection against stenosis, andhence coronary risk, by way of removal of excess cholesterol(transporting it to the liver for disposal).

A third group of carrier molecules, very low-density lipoproteins(VLDLs) are converted to LDLs following the delivery of triglycerides tothe muscles and adipose tissue. Triglycerides are a mixture of fattyacids and glycerol and are the major components of lipids circulating inblood. Like cholesterol, triglycerides are substances that are foundendogenously in the bloodstream, and may be deposited in adipose tissue.Triglycerides contain high-energy fatty acids which provide much of thefuel needed for normal cellular function. However, an excessive amountof triglycerides, or VLDLs, in the bloodstream can result in similarproblems to those associated with high cholesterol and LDL levels, aswell as obesity and diabetes.

Thus, levels of HDLs, LDLs, total cholesterol and triglycerides are allkey indicators in determining the risk of atherosclerosis and associatedcardiovascular disorders, such as coronary artery diseases (e.g. anginapectoris, myocardial infarction, etc.), stroke (includingcerebro-vascular accident and transient ischaemic attack), peripheralarterial occlusive disease, obesity and diabetes. Patients with highoverall cholesterol and/or triglycerides levels are at a significantrisk, irrespective of whether or not they also have a favourable HDLlevel. Patients with normal cholesterol levels but low HDL levels arealso at increased risk. Recently, it has also been noted that the levelof risk of cardiovascular disease associated with high levels ofapolipoprotein B (ApoB; which carries lipids in VLDLs and LDLs), and/orlow levels of apolipoprotein A-I (ApoA-I; which carries lipids in HDLs),is extremely high.

There are numerous factors that influence cholesterol and triglyceridelevels, including diet, age, weight, gender, genetics, diseases (such asdiabetes) and lifestyle.

Positive changes in relation to diet, lifestyle and exercise are ofteninsufficient to decrease the risk of cardiovascular problems. In suchinstances, cholesterol- and/or triglyceride-lowering medication may beprescribed.

Drugs that reduce LDL levels in serum can prevent or reduce the build-upof artery blocking plaques, and can reduce the risk of plaque ruptureand associated thrombo-embolic complications. There are several types ofdrugs that can help reduce blood cholesterol levels. The most commonlyprescribed are the statins, HMG-CoA reductase inhibitors, such aslovastatin, pravastatin, fluvastatin, simvastatin, atorvastatin,pitavastatin and rosuvastatin (e.g. rosuvastatin-calcium). These drugsprevent directly the formation of cholesterol in the liver and thusreduce the risk of cardiovascular disease. Other prescribed drugcategories include resins (such as cholestyramine and colestipol), whichact by binding bile acids, so causing the liver to produce more of thelatter, and using up cholesterol in the process. Further, the B vitaminniacin has been reported at high doses to lower triglycerides and LDLlevels in addition to increasing HDL levels. Fibrates (such asgemfibrozil and fenofibrate) are known to lower triglycerides and canincrease HDL levels.

However, some of these drugs are known to have side effects, includingliver damage. Hence, there is a need for alternative and/or moreeffective drugs for use in cholesterol-lowering therapy.

The early development of low molecular weight inhibitors of thrombin hasbeen described by Claesson in Blood Coagul. Fibrinol. (1994) 5, 411. Lowmolecular weight thrombin inhibitors (and prodrugs thereof) have beendescribed more recently in U.S. Pat. No. 4,346,078; International PatentApplications WO 93/11152, WO 93/18060, WO 93/05069, WO 94/20467, WO94/29336, WO 95/35309, WO 95/23609, WO 96/03374, WO 96/06832, WO96/06849, WO 96/17860, WO 96/24609, WO 96/25426, WO 96/32110, WO97/01338, WO 97/02284, WO 97/15190, WO 97/23499, WO 97/30708, WO97/40024, WO 97/46577, WO 98/06740, WO 97/49404, WO 97/11693, WO97/24135, WO 97/47299, WO 98/01422, WO 98/57932, WO 99/29664, WO98/06741, WO 99/37668, WO 99/37611, WO 98/37075, WO 99/00371, WO99/28297, WO 99/29670, WO 99/40072, WO 99/54313, WO 96/31504, WO00/01704, WO 00/08014, WO 00/35859, WO 00/35869, WO 00/42059, WO00/61577, WO 00/61608, WO 00/61609, WO 01/87879, WO 02/14270, WO02/44145, WO 03/000653, WO 04/000818 and WO 04/014894; and EuropeanPatent Applications 648 780, 468 231, 559 046, 641 779, 185 390, 526877, 542 525, 195 212, 362 002, 364 344, 530 167, 293 881, 686 642, 669317, 601 459 and 623 596.

In particular, international patent application WO 94/29336 discloses agroup of compounds, including HOOC—CH₂—(R)Cgl-(S)Aze-Pab-H (in which Cglrepresents cyclohexylglycyl, Aze represents azetidine-2-carboxyl andPab-H represents 4-amidinobenzylamino), which is also known asmelagatran (see Example 1 of WO 94/29336). International PatentApplication WO 97/23499 discloses prodrugs of inter alia melagatran.

More recently, international patent application WO 02/44145 disclosesα-hydroxy acid-based low molecular weight thrombin inhibitors andprodrugs thereof.

To the applicant's knowledge, none of the above-mentioned documentsdisclose or suggest the direct use a low molecular weight thrombininhibitor or a prodrug thereof in cholesterol-lowering therapy and/ormodifications of lipid (including triglyceride), lipoprotein, orapolipoprotein, profiles.

DISCLOSURE OF THE INVENTION

We have found, surprisingly, that administration of a low molecularweight thrombin inhibitor may give rise to reduced levels of lipids,such as total cholesterol, LDLs (i.e. LDL-cholesterol) and triglyceridesin the bloodstream, in addition to increasing HDL (i.e. HDL-cholesterol)levels.

According to a first aspect of the invention there is provided the useof a low molecular weight thrombin inhibitor, or a pharmaceuticallyacceptable derivative thereof, for the manufacture of a medicament foruse in cholesterol-lowering therapy.

When employed in the context of the present invention and disclosure,the term “cholesterol-lowering therapy” includes any therapy thatresults in beneficial modifications of serum profiles of totalcholesterol, lipids (including triglycerides), lipoproteins orapolipoproteins, and will thus be understood to encompass the terms“lipid-modifying therapy” and “lipid-(and triglyceride-) loweringtherapy”, as well as the treatment of hyperlipidaemias (i.e. theelevation of lipids in the bloodstream), including hypercholesterolaemia(high cholesterol levels in the blood; including primary and secondary(combined) hypercholesterolaemia), hyperlipoproteinemia (elevated plasmalipoproteins levels) and/or hypertriglyceridemia (high triglyceridelevels in the blood). The term will thus be understood to include typesI, II (IIa and IIb), III, IV and/or V hyperlipoproteinaemia, as well assecondary hypemmiglyceridaemia and/or familial lecithin cholesterolacyltransferase deficiency, but in principle includes any treatment of apatient which results in a decrease in serum levels of cholesterol,LDLs, VLDLs, triglycerides and/or ApoB, and/or an increase in serumlevels of HDLs and/or ApoA-I.

According to a second aspect of the invention there is provided acholesterol-lowering therapy method, which method comprises theadministration of a low molecular weight thrombin inhibitor, or apharmaceutically acceptable derivative thereof, to a patient in need ofsuch therapy.

For the avoidance of doubt, in the context of this disclosure, the terms“treatment”, “therapy” and “therapy method” include the therapeuticand/or prophylactic treatment of patients in need of modifications ofcholesterol, lipid (including triglyceride), lipoprotein and/orapolipoprotein profiles.

“Pharmaceutically acceptable derivatives” of thrombin inhibitorsincludes salts (e.g. pharmaceutically-acceptable non-toxic organic orinorganic acid addition salts) and solvates. It will be appreciated thatthe term further includes derivatives that have, or provide for, thesame biological function and/or activity as any relevant inhibitor.Thus, for the purposes of this invention, the term also includesprodrugs of thrombin inhibitors.

The term “low molecular weight thrombin inhibitor” will be understood bythose skilled in the art. The term may also be understood to include anycomposition of matter (e.g. chemical compound) which inhibits thrombinto an experimentally determinable degree in in vivo and/or in in vitrotests, and which possesses a molecular weight of below 2,000, preferablybelow 1,000.

Preferred low molecular weight thrombin inhibitors include low molecularweight peptide-based, amino acid-based, and/or peptide analogue-based,thrombin inhibitors.

The term “low molecular weight peptide-based, amino acid-based, and/orpeptide analogue-based, thrombin inhibitors” will be well understood byone skilled in the art to include low molecular weight thrombininhibitors with one to four peptide linkages, and includes thosedescribed in the review paper by Claesson in Blood Coagul. Fibrin.(1994) 5, 411, as well as those disclosed in U.S. Pat. No. 4,346,078;International Patent Applications WO 93/11152, WO 93/18060, WO 93/05069,WO 94/20467, WO 94/29336, WO 95/35309, WO 95/23609, WO 96/03374, WO96/06832, WO 96/06849, WO 96/17860, WO 96/24609, WO 96/25426, WO96/32110, WO 97/01338, WO 97/02284, WO 97/15190, WO 97/30708, WO97/40024, WO 97/46577, WO 98/06740, WO 97/49404, WO 97/11693, WO97/24135, WO 97/47299, WO 98/01422, WO 98/57932, WO 99/29664, WO98/06741, WO 99/37668, WO 99/37611, WO 98/37075, WO 99/00371, WO99/28297, WO 99/29670, WO 99/40072, WO 99/54313, WO 96/31504, WO00/01704, WO 00/08014, WO 00/35859, WO 00/35869, WO 00/42059, WO00/61577, WO 00/61608, WO 00/61609, WO 01/87879, WO 02/14270, WO02/44145, WO 03/000653 and WO 04/000818; and European PatentApplications 648 780, 468 231, 559 046, 641 779, 185 390, 526 877, 542525, 195 212, 362 002, 364 344, 530 167, 293 881, 686 642, 669 317, 601459 and 623 596, the specific and generic disclosures in all of whichdocuments are hereby incorporated by reference.

Preferred low molecular weight peptide-based thrombin inhibitors includethose described generically and specifically in international patentapplication WO 98/37075, including the compound that is the subject ofclaim 8 of that application as published(1-methyl-2-[N-(4-amidinophenyl)aminomethyl]-benzimidazol-5-yl-carboxylicacid, N-(2-pyridyl)-N-(2-hydroxycarbonyl-ethyl)amide) and prodrugsthereof, such as the compound that is the subject of claim 10 of thatapplication as published(1-methyl-2-[N-[4-(N-n-hexyloxycarbonylamidino)phenyl]aminomethyl]benzimidazol-5-yl-carboxylicacid, N-(2-pyridyl)-N-(2-ethoxycarbonylethyl)amide) and those describedin international patent application WO 04/014894.

Preferred low molecular weight peptide-based thrombin inhibitors alsoinclude HOOC—CH₂—(R)Cha-Pic-Nag-H (wherein Cha representscyclohexylalanine, Pic represents (S)-pipecolinic acid and Nagrepresents noragmatine; known as inogatran; see International PatentApplication WO 93/11152) and, especially, HOOC—CH₂—(R)Cgl-(S)Aze-Pab-H(known as melagatran; see above and International Patent Application WO94/29336).

Further thrombin inhibitors include those of the formula I,

whereinR^(a) represents —OH or —CH₂OH;R¹ represents at least one optional halo substituent;R² represents one or two C₁₋₃ alkoxy substituents, the alkyl parts ofwhich substituents are themselves substituted with one or more fluorosubstituents (i.e. R² represents one or two fluoroalkoxy(C₁-3) groups);Y represents —CH₂— or —(CH₂)₂—; andR³ represents a structural fragment of formula I(i) or I(ii):

whereinR⁴ represents H or one or more fluoro substituents; andone or two of X₁, X₂, X₃ and X₄ represent —N— and the others represent—CH—.

Preferred compounds of formula I include:(a) Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)-(S)Aze-Pab:

(b) Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF):

(c) Ph(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab:

The term “prodrug” of a low molecular weight thrombin inhibitor includesany compound that, following oral or parenteral administration, ismetabolised in vivo to form a low molecular weight thrombin inhibitor(as defined herein), in an experimentally-detectable amount, and withina predetermined time (e.g. within a dosing interval of between 6 and 24hours (i.e. once to four times daily)). For the avoidance of doubt, theterm “parenteral” administration includes all forms of administrationother than oral administration.

Prodrugs of the thrombin inhibitor melagatran that may be mentionedinclude those disclosed in international patent application WO 97/23499.Preferred prodrugs are those of the formulaR¹O₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH (see the list of abbreviations above orin WO 97/23499), wherein R¹ represents C₁₋₁₀ alkyl or benzyl, such aslinear or branched C₁₋₆ alkyl (e.g. C₁₋₄ alkyl, especially methyl,propyl and, particularly, ethyl) and the OH group replaces one of theamidino hydrogens in Pab.

Preferred prodrugs of compounds of formula I that may be mentionedinclude those of formula Ia,

wherein R^(3a) represents a structural fragment of formula I(iii) orI(iv):

wherein R⁵ represents OR⁶ or C(O)OR⁷;R⁶ represents H, C₁₋₁₀ alkyl, C₁₋₃ alkylaryl or C₁₋₃ alkyloxyaryl (thealkyl parts of which latter two groups are optionally interrupted by oneor more oxygen atoms, and the aryl parts of which latter two groups areoptionally substituted by one or more substituents selected from halo,phenyl, methyl or methoxy, which latter three groups are also optionallysubstituted by one or more halo substituents);R⁷ represents C₁₋₁₀ alkyl (which latter group is optionally interruptedby one or more oxygen atoms), or C₁₋₃ alkylaryl or C₁₋₃ alkyloxyaryl(the alkyl parts of which latter two groups are optionally interruptedby one or more oxygen atoms, and the aryl parts of which latter twogroups are optionally substituted by one or more substituents selectedfrom halo, phenyl, methyl or methoxy, which latter three groups are alsooptionally substituted by one or more halo substituents); andR^(a), R¹, R², Y, R⁴, X₁, X₂, X₃ and X⁴ are as hereinbefore defined.

Preferred prodrugs of compounds of formula I are methoxyamidine prodrugsthereof. Hence preferred compounds of formula Ia include:(i) Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe):

(ii) Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)-(S)Aze-Pab(2,6-diF)(OMe):

(iii) Ph(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)-(S)Aze-Pab(OMe):

Compounds of formulae I and Ia may be made in accordance with techniquesdescribed in international patent application WO 02/44145.

In accordance with the invention, thrombin inhibitors and derivativesthereof may be administered orally, intravenously, subcutaneously,buccally, rectally, dermally, nasally, tracheally, bronchially,topically, by any other parenteral route, or via inhalation, in the formof a pharmaceutical preparation comprising the thrombin inhibitor orprodrug in a pharmaceutically acceptable dosage form. Depending on thedisorder, and the patient, to be treated, as well as the route ofadministration, the compositions may be administered at varying doses.

Preferred modes of delivery are systemic. For melagatran and derivativesthereof, preferred modes of administration are parenteral, morepreferably intravenous, and especially subcutaneous. For prodrugs ofmelagatran and compounds of formula Ia, preferred modes ofadministration are oral.

In the therapeutic treatment of manmals, and especially humans, thrombininhibitors, prodrugs of thrombin inhibitors, and derivatives of eitherwill generally be administered as a pharmaceutical formulation inadmixture with a pharmaceutically acceptable adjuvant, diluent orcarrier, which may be selected with due regard to the intended route ofadministration and standard pharmaceutical practice.

Suitable formulations for use in administering thrombin inhibitors areknown in the art, and include those known from U.S. Pat. No. 4,346,078;International Patent Applications WO 93/11152, WO 93/18060, WO 93/05069,WO 94/20467, WO 94/29336, WO 95/35309, WO 95/23609, WO 96/03374, WO96/06832, WO 96/06849, WO 96/17860, WO 96/24609, WO 96/25426, WO96/32110, WO 97/01338, WO 97/02284, WO 97/15190, WO 97/30708, WO97/40024, WO 97/46577, WO 98/06740, WO 97/49404, WO 97/11693, WO97/24135, WO 97/47299, WO 98/01422, WO 98/57932, WO 99/29664, WO98/06741, WO 99/37668, WO 99/37611, WO 98/37075, WO 99/00371, WO99/28297, WO 99/29670, WO 99/40072, WO 99/54313, WO 96/31504, WO00/01704, WO 00/08014, WO 00/35859, WO 00/35869, WO 00/42059, WO00/61577, WO 00/61608, WO 00/61609, WO 01/87879, WO 02/14270, WO02/44145, WO 03/000653 and WO 04/000818; and European PatentApplications 648 780, 468 231, 559 046, 641 779, 185 390, 526 877, 542525, 195 212, 362 002, 364 344, 530 167, 293 881, 686 642, 669 317, 601459 and 623 596, the disclosures in all of which documents are herebyincorporated by reference.

Suitable formulations for use with melagatran, derivatives and prodrugsthereof are described in the literature, for example as described ininter alia international patent applications WO 94/29336, WO 96/14084,WO 96/16671, WO 97/23499, WO 97/39770, WO 97/45138, WO 98/16252, WO99/27912, WO 99/27913, WO 00/12043 and WO 00/13671, the disclosures inwhich documents are hereby incorporated by reference. Otherwise, thepreparation of suitable formulations may be achieved non-inventively bythe skilled person using routine techniques.

The amounts of thrombin inhibitor or derivative in the formulation willdepend on the severity of the condition, and on the patient, to betreated, as well as the compound(s) which is/are employed, but may bedetermined non-inventively by the skilled person.

Suitable doses of thrombin inhibitors and derivatives thereof in thetherapeutic and/or prophylactic treatment of mammalian, especiallyhuman, patients may be determined routinely by the medical practitioneror other skilled person, and include the respective doses discussed inthe relevant prior art documents that are mentioned hereinbefore, therelevant disclosures in which are hereby incorporated by reference.

In the case of melagatran, suitable doses of active compound, prodrugsand derivatives thereof, in the therapeutic and/or prophylactictreatment of mammalian, especially human, patients include those whichgive a mean plasma concentration of up to 5 μmol/L, for example in therange 0.001 to 5 μmol/L over the course of treatment of the relevantcondition. Suitable doses may thus be in the range 0.1 mg once daily to25 mg three times daily, and/or up to 100 mg infused parenterally over a24 hour period, for melagatran, and in the range 0.1 mg once daily to100 mg three times daily for prodrugs of melagatran (see also thespecific doses mentioned hereinafter for the prodrug of melagatran,ximelagatran).

In the case of compounds of formulae I and Ia, suitable daily doses ofcompounds in the therapeutic treatment of humans are about 0.001-100mg/kg body weight at peroral administration and 0.001-50 mg/kg bodyweight at parenteral administration.

In any event, the physician, or the skilled person, will be able todetermine the actual dosage which will be most suitable for anindividual patient, which is likely to vary with the severity of thecondition that is to be treated, as well as the age, weight, sex andresponse of the particular patient to be treated. The above-mentioneddosages are exemplary of the average case; there can, of course, beindividual instances where higher or lower dosage ranges are merited,and such are within the scope of this invention.

Low molecular weight thrombin inhibitors may be employed in the methoddescribed herein by way of co-administration along with othercholesterol-lowering, or lipid-lowering/modifying, drugs/therapies thatare mentioned hereinbefore, such as the statins (HMG-CoA reductaseinhibitors) and particularly any one of those statins specificallymentioned hereinbefore, in combination therapy.

According to a further aspect of the invention, there is provided acombination product comprising:

-   (A) a low molecular weight thrombin inhibitor, or a    pharmaceutically-acceptable derivative thereof; and-   (B) another cholesterol-lowering, or lipid-lowering/modifying,    therapeutic agent,    wherein each of components (A) and (B) is formulated in admixture    with a pharmaceutically-acceptable adjuvant, diluent or carrier.

Such combination products provide for the administration of lowmolecular weight thrombin inhibitor/derivative in conjunction with theother therapeutic agent, and may thus be presented either as separateformulations, wherein at least one of those formulations comprisesthrombin inhibitor/derivative, and at least one comprises the othertherapeutic agent, or may be presented (i.e. formulated) as a combinedpreparation (i.e. presented as a single formulation including thrombininhibitor/derivative and the other therapeutic agent).

Thus, there is further provided:

(1) a pharmaceutical formulation including a low molecular weightthrombin inhibitor, or a pharmaceutically-acceptable derivative thereof;another cholesterol-lowering, or lipid lowering/modifying, therapeuticagent; and a pharmaceutically-acceptable adjuvant, diluent or carrier;and

(2) a kit of parts comprising components:

-   (a) a pharmaceutical formulation including a low molecular weight    thrombin inhibitor, or a pharmaceutically-acceptable derivative    thereof, in admixture with a pharmaceutically-acceptable adjuvant,    diluent or carrier; and-   (b) a pharmaceutical formulation including another    cholesterol-lowering, or lipid lowering/modifying, therapeutic agent    in admixture with a pharmaceutically-acceptable adjuvant, diluent or    carrier,    which components (a) and (b) are each provided in a form that is    suitable for administration in conjunction with the other.

The method described herein may have the advantage that, incholesterol-lowering therapy, it may be more convenient for thephysician and/or patient than, be more efficacious than, be less toxicthan, have a broader range of activity than, be more potent than,produce fewer side effects than, or that it may have other usefulpharmacological properties over, similar methods (treatments) known inthe prior art for use in such therapy.

The invention is illustrated, but in no way limited, by the followingexamples, in which:

FIG. 1 illustrates the difference in mean values (with 95% ConfidenceIntervals) of total cholesterol levels in serum as between patientsreceiving ximelagatran (36 mg bid) or warfarin (INR levels between 2 and3) during the course of a clinical trial over a 21 month period.

FIG. 2 illustrates the difference in mean values (with 95% ConfidenceIntervals) of levels of total triglycerides in serum as between patientsreceiving ximelagatran (36 mg bid) or warfarin (INR levels between 2 and3) during the course of a clinical trial over a 21 month period.

FIG. 3 illustrates the difference in mean values (with 95% ConfidenceIntervals) of LDL (i.e. LDL-cholesterol) levels in serum as betweenpatients receiving ximelagatran (36 mg bid) or warfarin (INR levelsbetween 2 and 3) during the course of a clinical trial over a 21 monthperiod.

FIG. 4 illustrates the difference in mean values (with 95% ConfidenceIntervals) of HDL (i.e. HDL-cholesterol) levels in serum as betweenpatients receiving ximelagatran (36 mg bid) or warfarin (INR levelsbetween 2 and 3) during the course of a clinical trial over a 21 monthperiod.

EXAMPLE 1

Lipid Measurements in Patients Undergoing Thrombin Inhibition Therapy ina Clinical Trial

A large-scale Phase III clinical trial was set up to establish theefficacy of the study compound EtO₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH(ximelagatran; see Example 17 of international patent application WO97/23499) in the prevention of stroke in patients with non-valvularatrial fibrillation, as compared to the current frontline treatment forthis indication, warfarin.

Ximelagatran is a prodrug of the low molecular weight thrombininhibitor, melagatran (see Example 1 of international patent applicationWO 94/29226).

The clinical trial protocol was similar to that described ininternational patent application WO 02/36157, with the following majordifferences:

-   (a) the study objective was to show that the efficacy of    ximelagatran is non-inferior to that of dose-adjusted warfarin,    aiming for an INR 2.0-3.0 (with INR measurements taken at least    every 28±3 days), in the prevention of all strokes (fatal and    non-fatal) and systemic embolic events in patients with chronic    non-valvular atrial fibrillation;-   (b) the dosage of ximelagatran was fixed at 36 mg bid;-   (c) in the exclusion criteria, subjects who had experienced stroke    within the previous 30 days or transient ischaemic attack within the    previous 3 days were excluded (as opposed to 2 years in the study    described in WO 02/36157);-   (d) the duration of treatment was long term (between 12 and 26    months); and-   (e) the total number of patients in the trial was 3407 (as opposed    to 220 in the study described in WO 02/36157). The study was a    multicentre, multinational, IVRS-randomised, open-label,    parallel-group study carried out across approximately 300 centres in    approximately 25 countries.

Over the first 21 months of the trial, blood samples were taken from allpatients in a standard manner at the following intervals: beforehand,and at, 1, 1.5, 2, 3, 4, 5, 6, 8, 10, 12, 15, 18, and 21, months. Thesesamples were routinely analysed for total cholesterol, LDL (i.e.LDL-cholesterol), HDL (i.e. HDL-cholesterol), and total triglycerides,content using standard techniques for the detection of these lipids inserum.

A comparison was made between patients on ximelagatran (n=1704) andwarfarin (n=1703). When the raw data were analysed, an unexpectedstatistically-significant difference in favour of the ximelagatran groupwas observed. As of the second month of treatment, marked meandifferences were observed for cholesterol, triglycerides and LDL serumconcentrations (consistently significantly lower in the ximelagtrangroup over the entire 21 month period), and for the HDL serumconcentration (consistently significantly higher in the ximelagtrangroup over the entire 21 month period), as illustrated in FIGS. 1 to 4,respectively.

These data clearly demonstrate the potential utility of melagatran andderivatives thereof (e.g. prodrugs, such as ximelagtran), as well as,potentially, low molecular weight thrombin inhibitors andderivatives/prodrugs thereof, in cholesterol-lowering therapy.

1-13. (canceled)
 14. A cholesterol-lowering therapy method, which methodcomprises the administration of melagatran inhibitor, or apharmaceutically acceptable derivative thereof to a patient in need ofsuch therapy.
 15. (canceled)
 16. The method of claim 14 wherein thetherapy/treatment results in a decrease in serum levels of cholesterol,low-density lipoproteins, very low-density lipoproteins, triglyceridesand/or apolipoprotein B; and/or an increase in serum levels ofhigh-density lipoproteins and/or apolipoprotein A-I.
 17. (canceled) 18.The method of claim 14, wherein the method comprises administering aprodrug of melagatran.
 19. The method of claim 18, wherein the methodcomprises delivering a prodrug is of the formula:R¹O₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein R¹ represents linear or branchedC₁₋₆ alkyl and the OH group replaces one of the amidino hydrogens inPab.
 20. The method of claim 19, wherein R¹ represents methyl, ethyl, orpropyl.
 21. The method of claim 20, wherein R¹ represents ethyl.
 22. Acholesterol-lowering therapy method, which method comprises theadministration of a thrombin inhibitor of formula I,

wherein R^(a) represents —OH or —CH₂OH; R¹ represents at least oneoptional halo substituent; R² represents one or two C₁₋₃ alkoxysubstituents, the alkyl parts of which substituents are themselvessubstituted with one or more fluoro substituents; Y represents —CH₂— or—(CH₂)₂—; and R³ represents a structural fragment of formula I(i) orI(ii):

wherein R⁴ represents H or one or more fluoro substituents; and one ortwo of X₁, X₂, X₃, and X₄ represent —N— and the others represent —CH—,or a pharmaceutically acceptable derivative thereof, to a patient inneed of such therapy.
 23. The method of claim 22, wherein the thrombininhibitor or derivative is: Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab;Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab.
 24. The method of claim22, wherein the the thrombin inhibitor or derivative is a prodrug of athrombin inhibitor.
 25. The method of claim 24, wherein the prodrug isof formula Ia,

wherein R^(3a) represents a structural fragment of formula I(iii) orI(iv):

wherein R⁵ represents OR⁶ or C(O)OR⁷; R⁶ represents H, C₁₋₁₀ alkyl, C₁₋₃alkylaryl, or C₁₋₃ alkyloxyaryl, the alkyl parts of which latter twogroups are optionally interrupted by one or more oxygen atoms, and thearyl parts of which latter two groups are optionally substituted by oneor more substituents selected from halo, phenyl, methyl or methoxy,which latter three groups are also optionally substituted by one or morehalo substituents; and R⁷ represents C₁₋₁₀ alkyl, optionally interruptedby one or more oxygen atoms; C₁₋₃ alkylaryl; or C₁₋₃ alkyloxyaryl, thealkyl parts of which latter two groups are optionally interrupted by oneor more oxygen atoms, and the aryl parts of which latter two groups areoptionally substituted by one or more substituents selected from halo,phenyl, methyl or methoxy, which latter three groups are also optionallysubstituted by one or more halo substituents.
 26. The method of claim25, wherein the prodrug is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe);Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF)(OMe); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe).
 27. A combinationproduct comprising: (A) a low molecular weight thrombin inhibitor, or apharmaceutically-acceptable derivative thereof; and (B) anothercholesterol-lowering, or lipid-lowering/modifying, therapeutic agent,wherein each of components (A) and (B) is formulated in admixture with apharmaceutically-acceptable adjuvant, diluent or carrier. 28-29.(canceled)
 30. The combination product of claim 27, wherein the thrombininhibitor or derivative is melagatran.
 31. A The combination product ofclaim 27, wherein the thrombin inhibitor or derivative is a prodrug ofmelagatran.
 32. The combination product of claim 31, wherein the prodrugis of the formula:R¹O₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein R¹ represents linear or branchedC₁₋₆ alkyl and the OH group replaces one of the amidino hydrogens inPab.
 33. The combination product of claim 32, wherein R¹ representsmethyl, ethyl, or propyl.
 34. The combination product in of claim 33,wherein R¹ represents ethyl.
 35. The combination product of claim 27,wherein the thrombin inhibitor or derivative is a compound of formula I,

wherein R^(a) represents —OH or —CH₂OH; R¹ represents at least oneoptional halo substituent; R² represents one or two C₁₋₃ alkoxysubstituents, the alkyl parts of which substituents are themselvessubstituted with one or more fluoro substituents; Y represents —CH₂— or—(CH₂)₂—; and R³ represents a structural fragment of formula I(i) orI(ii):

wherein R⁴ represents H or one or more fluoro substituents; and one ortwo of X₁, X₂, X₃, and X₄ represent —N— and the others represent —CH—.36. The combination product of claim 27, wherein the thrombin inhibitoror derivative is: Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab;Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab.
 37. The combinationproduct of claim 27, wherein the thrombin inhibitor or derivative is aprodrug of a thrombin inhibitor.
 38. The combination product of claim27, wherein the thrombin inhibitor or derivative is a prodrug is offormula Ia,

wherein R^(3a) represents a structural fragment of formula I(iii) orI(iv):

wherein R⁵ represents OR⁶ or C(O)OR⁷; R⁶ represents H, C₁₋₁₀ alkyl, C₁₋₃alkylaryl or C₁₋₃ alkyloxyaryl, the alkyl parts of which latter twogroups are optionally interrupted by one or more oxygen atoms, and thearyl parts of which latter two groups are optionally substituted by oneor more substituents selected from halo, phenyl, methyl or methoxy,which latter three groups are also optionally substituted by one or morehalo substituents; and R⁷ represents C₁₋₁₀ alkyl, which latter group isoptionally interrupted by one or more oxygen atoms; C₁₋₃ alkylaryl; orC₁₋₃ alkyloxyaryl, the alkyl parts of which latter two groups areoptionally interrupted by one or more oxygen atoms, and the aryl partsof which latter two groups are optionally substituted by one or moresubstituents selected from halo, phenyl, methyl or methoxy, which latterthree groups are also optionally substituted by one or more halosubstituents.
 39. The combination product of claim 27, wherein thethrombin inhibitor or derivative is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe);Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF)(OMe); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe).
 40. The combinationproduct of claim 27, wherein the other therapeutic agent is a statin.41. The combination product of claim 27, wherein the other therapeuticagent is lovastatin, pravastatin, fluvastatin, simvastatin,atorvastatin, pitavastatin, or rosuvastatin. 42-44. (canceled)
 45. Themethod of claim 22, wherein the therapy/treatment results in a decreasein serum levels of cholesterol, low-density lipoproteins, verylow-density lipoproteins, triglycerides and/or apolipoprotein B; and/oran increase in serum levels of high-density lipoproteins and/orapolipoprotein A-I.
 46. The combination product of claim 27, wherein thethrombin inhibitor or derivative is of the formula:R¹O₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein R¹ represents linear or branchedC₁₋₆ alkyl and the OH group replaces one of the amidino hydrogens inPab, and the other therapeutic agent is a statin.
 47. The combinationproduct of claim 27, wherein the thrombin inhibitor or derivative is ofthe formula:R¹O₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein R¹ represents linear or branchedC₁₋₆ alkyl and the OH group replaces one of the amidino hydrogens inPab, and the other therapeutic agent is lovastatin, pravastatin,fluvastatin, simvastatin, atorvastatin, pitavastatin, or rosuvastatin.48. The combination product of claim 27, wherein the thrombin inhibitoror derivative is of the formula:EtO₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein Et represents ethyl and the OHgroup replaces one of the amidino hydrogens in Pab, and the othertherapeutic agent is a statin
 49. The combination product of claim 27,wherein the thrombin inhibitor or derivative is of the formula:EtO₂C—CH₂—(R)Cgl-(S)Aze-Pab-OH, wherein Et represents ethyl and the OHgroup replaces one of the amidino hydrogens in Pab, and the othertherapeutic agent is lovastatin, pravastatin, fluvastatin, simvastatin,atorvastatin, pitavastatin, or rosuvastatin.
 50. The combination productof claim 27, wherein the thrombin inhibitor or derivative is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab;Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab, and the other therapeuticagent is a statin.
 51. The combination product of claim 27, wherein thethrombin inhibitor or derivative is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe);Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF)(OMe); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe), and the othertherapeutic agent is a statin.
 52. The combination product of claim 27,wherein the thrombin inhibitor or derivative is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab;Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab, and the other therapeuticagent is lovastatin, pravastatin, fluvastatin, simvastatin,atorvastatin, pitavastatin, or rosuvastatin.
 53. The combination productof claim 27, wherein the thrombin inhibitor or derivative is:Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe);Ph(3-Cl)(5-OCHF₂)—(R)CH(OH)C(O)—(S)Aze-Pab(2,6-diF)(OMe); orPh(3-Cl)(5-OCH₂CH₂F)—(R)CH(OH)C(O)—(S)Aze-Pab(OMe), and the othertherapeutic agent is lovastatin, pravastatin, fluvastatin, simvastatin,atorvastatin, pitavastatin, or rosuvastatin.